Cyclic nucleotide phosphodiesterase (PDE) enzymes are a superfamily of intracellular enzymes. In humans, 21 PDE genes are known, belonging to 11 PDE families. However, many more than 21 mRNA and protein products are known to be transcribed from these 21 genes, as alternative transcriptional start sites and alternative splicing of mRNA precursor molecules exist. Hence, the 11 PDE families encompass over 60 isoforms.
PDEs degrade cyclic nucleotides, in particular cyclic adenosine 3′,5′-monophosphate (cAMP) and cyclic guanosine 3′,5′-monophosphate (cGMP). cAMP and cGMP are intracellular second messengers playing a key role in mediating cellular responses to various hormones and neurotransmitters. The levels of cAMP and cGMP are tightly regulated by their synthesis (by the enzymes adenylate and guanylate cyclase) as well as their degradadtion (by the PDEs). PDEs render the ability to generate and shape cAMP or cGMP gradients within the cell.
The 11 different PDE families have different affinities for cAMP and cGMP, and each family is therefore involved in the degradation of one or both of these cyclic nucleotides. Any single cell type can express several different PDEs and the nature and intracellular localization of these PDEs is an important regulator of local concentrations of cAMP and cGMP within the cell. The regulation of intracellular cyclic nucleotide levels by PDEs is a complex interplay between the multiple PDE isoforms. Although there may be some redundancy between the different PDE isoforms, it is known that most of the different PDE isoforms have specific physiological roles.
The 11 PDE families are named PDE1 through PDE11, the number referring to the particular gene family. To indicate the gene within the family, a capital letter can be added to this name. Further numbers specifying the variant can then be further added to the name. Furthermore, often a species code is added preceding the name, e.g. Hs stands for Homo sapiens. An example of a PDE name according to this nomenclature would be HsPDE1A2. In an alternative annotation system, PDEs are written in lower case italic letters when referring to a gene, and in non italicized capital letters when referring to a protein, and a letter “v” is added preceding the variant number. An example of a PDE gene name and the corresponding PDE protein name according to this nomenclature would be Pde1a_v2 and PDE1A_V2, respectively.
When analyzing crystallographic data, PDE catalytic domains have been found to consist of a compact arrangement of 16 α-helices arranged in three subdomains. The active site is located at the intersection of these three subdomains. Of the 16 invariant amino acids across the 21 PDE genes, 11 are located in the active site. Two metal ions, in particular magnesium and/or zinc, are coordinated each by six residues found at the bottom of the active site. These residues are located on each of the three subdomains. As two metal ions are needed, a binuclear catalytic mechanism for cleaving the cyclic phosphate group of cAMP or cGMP has been suggested. To accommodate the substrate, conserved aromatic residues at the roof of the active site enable a stacking arrangement with the cyclic nucleotide, a hydrophobic residue at the bottom of the active site enables hydrophobic interaction, and further hydrogen bonds are formed with the conserved active site glutamine, finally the phosphate group interacts with the two metal ions directly or via a water molecule. The conserved active site glutamine, which is also named glutamine switch, is a key determinant of cyclic nucleotide specificity. The glutamine switch stabilizes the binding of the cyclic nucleotide purine ring in the active site. To accommodate the binding of both cAMP and cGMP, free rotation of this conserved glutamine residue is required. In PDEs which are selective for one of the cyclic nucleotides, the free rotation of the conserved glutamine residue is constrained by neighboring residues into an orientation favoring cAMP binding only, or into an orientation favoring cGMP binding only.
The PDE4 family, encoded by four genes (PDE4A, PDE4B, PDE4C, and PDE4D) and containing over 20 identified isoforms, is the largest of the 11 PDE families. PDE4 is a cAMP-specific phosphodiesterase, hence its original name cAMP-PDE. PDE4 isoforms are distributed widely and can be found in most tissues and cell types, including the airways smooth muscle, brain, and cardiovascular tissues. Therefore, PDE4 inhibitors may target multiple cell types. Most PDE4 isoforms have tissue- and cell-type-specific expression. As PDE4 isoforms arise from differences in their N termini, which encode regulatory domains and phosphorylation sites, it will be appreciated that different isoforms are subject to different regulatory mechanisms. PDE4 isoforms have distinct cellular localizations as well.
PDE4 isoforms are omnipresent among proinflammatory and inflammatory cells, and immune cells, and PDE4 is the main cAMP-metabolizing enzyme in these cells. Raising intracellular cAMP levels within inflammatory cells, which can be achieved by inhibiting PDE4 activity, inhibits inflammatory cell function.
Diseases for which PDE4 inhibitors have been developed or are currently being developed include—but are not limited to—chronic obstructive pulmonary disease (COPD), asthma, dry eye disease, arthritis, psoriasis, atopic dermatitis, inflammatory bowel disease, and major depressive disorder. PDE4 inhibitors are also being investigated as memory-enhancing agents. In addition, PDE4 inhibitors are of potential interest for the treatment of fibrotic diseases, including Idiopathic Pulmonary Fibrosis (IPF).
COPD is a respiratory disease characterized by small airway fibrosis, mucus hypersecretion, and emphysema, resulting in a rapid decline in lung function. While the main genetic cause of COPD is α1 anti-trypsin deficiency, cigarette smoking is the major risk factor for COPD development. Inhaled cigarette smoke activates resident cells in the lungs including fibroblasts, epithelial cells, and alveolar macrophages to release cytokines, chemokines, and lipid mediators (including tumor necrosis factor α (TNF-α), interleukin-8 (IL-8), transforming growth factor β (TGF-β), and leukotriene B4 (LTB4)). As a result, inflammatory cells are recruited and activated to release a cocktail of proteases into the matrix compartment. This cocktail of proteases can provoke a complex remodeling process leading to alveolar wall destruction, mucus hypersecretion, and peribronchiolar fibrosis, ultimately resulting in COPD. Moreover, exposure to tobacco smoke can induce an imbalance in the oxidant versus antioxidant ratio in favor of oxidative stress. Inflammation can continue in patients long after smoking cessation. It is appreciated that targeting the inflammation and remodeling processes associated with COPD may slow down disease progression and potentially reverse the decline in lung function. Drugs frequently used to treat COPD include β2 agonists, anticholinergic agents, methylxanthines (such as the PDE inhibitor theophylline), and inhaled glucocorticosteroids. These drugs reduce exacerbations and symptoms of COPD, but may not reduce disease progression. Since the inflammation and remodeling process is driven by a number of mediators with overlapping roles, targeting individual mediators is unlikely to result in any therapeutic benefit. As PDE4 inhibitors can decrease/suppress the release of a wide range of inflammatory mediators, cytokines, chemokines, and proteases from cells implicated in COPD, their use in the treatment of COPD may be advantageous. In the clinic, PDE4 inhibitors, including roflumilast and cilomilast, have shown efficacy in COPD treatment.
Asthma is a chronic inflammatory disease of the airways characterized by variable and recurring symptoms, reversible airflow obstruction, and bronchospasm. Unlike COPD, the airway obstruction in asthma is usually reversible. If left untreated however, the chronic inflammation of the lungs can become irreversible obstruction due to airway remodeling. In contrast to emphysema, asthma affects the bronchi, not the alveoli. Asthma is caused by a combination of genetic and environmental factors. Bronchodilators are recommended for short-term relief of symptoms. Acute symptoms are usually treated with an inhaled short-acting β2 agonist. Inhaled corticosteroids or oral leukotriene antagonists are used for symptom prevention, while also avoidance of triggers might prevent asthma. For severe asthma exacerbation, oral glucocorticoids are added to the treatment. PDE4 inhibitors may have use in the inhibition of inflammatory cells involved in asthma. Moreover, as PDE4 inhibitors have been found to induce relaxation of isolated human bronchus, they might also possess bronchodilator activity, which is beneficial in the treatment of asthma. In the clinic, the PDE4 inhibitor roflumilast has been found to reduce the early asthmatic response to antigen challenge and to suppress the late asthmatic response. The late asthmatic response is viewed to represent the inflammatory component of airway disease. Moreover, roflumilast led to increased forced expiratory volume in 1 second (FEV1) and improved morning and evening peak expiratory flow.
Dry eye is a disease of the tears and the ocular surface, leading to ocular discomfort and blurred vision. The disease can be caused by numerous factors, but once established, inflammation of various ocular tissues propagates the disease as both cause and consequence of ocular surface damage. Artificial tears might improve symptoms, but the relief is temporary as the underlying inflammation persists. Given their anti-inflammatory properties, PDE4 inhibitors might reduce eye inflammation. Moreover, other agents that increase cAMP have been found to induce tear secretion, thus PDE4 inhibitors might induce tear secretion as well. Hence in the treatment of eye disease, PDE4 inhibitors might serve the dual role of inflammation reduction and induction of tear secretion.
In addition to COPD, asthma, and dry eye disease, other inflammatory diseases for which PDE4 inhibitors are being developed include inflammatory bowel disease, psoriasis, atopic dermatitis, and arthritis. These uses are based on the effect of decreased PDE4 activity to reduce inflammatory responses of multiple cell types. PDE4 inhibitors suppress the release of inflammatory mediators and immune cell infiltration, in particular, major dampening effects have been observed on neutrophil, monocyte, and T-lymphocyte function.
PDE4 inhibitors are also of interest for the treatment of fibrotic diseases. Fibrosis is the formation of excess fibrous connective tissue in an organ or tissue in a reparative or reactive process, following injury or long term inflammation. Fibrotic diseases are characterized by the accumulation of extracellular matrix together with distortion and disruption of tissue architecture. Among them, Pulmonary Fibrosis (PF) involves the overgrowth, hardening, and/or scarring of lung tissue due to excess collagen. The most common variation of this disease is Idiopathic (unknown cause) Pulmonary Fibrosis (IPF). IPF patients show decline in gas exchange and reduction in total lung volume. Chronic inflammation is a hallmark of PF, with increased amounts of inflammatory cells both in alveolar space and lung interstitium. Numbers of all inflammatory cells is dramatically increased in Bronchoalveolar lavage fluid (BALF) of PF patients, with boost in neutrophil and lymphocyte counts. While this increase in total BALF cell number is mostly accounted for by macrophages, maximal relative increase is observed for granulocytes and lymphocytes. Those changes are accompanied by elevated expression of cytokines such as TNF-α, IL-1β, IL-6, of growth factors and of matrix metalloproteases. As PDE4 inhibitors can modulate inflammatory cell function, it will be appreciated that such inhibitors are of interest for the treatment of fibrotic diseases such as IPF. Additionally, TGF-β has been shown to play a role in multiple fibrotic diseases, including pulmonary fibrosis, renal fibrosis, cardiac fibrosis and vascular fibrosis. TGF-β can potentially modulate cAMP by altering PGE2 metabolism. As a result of their effect on cAMP degradation, PDE4 inhibitors have been shown to functionally antagonize the profibrotic activity of fibroblasts stimulated by TGF-β1. It will therefore be appreciated that PDE4 inhibitors are of interest for the treatment of TGF-β-driven fibrotic diseases.
The prototypical PDE4 inhibitor is rolipram. This compound was originally developed for the treatment of major depressive disorder. It has been found that elevation of cAMP can enhance noradrenergic neurotransmission in the central nervous system. In the clinic, rolipram has been found to be an effective antidepressant, but its development has been stopped due to side effects. In addition, rolipram has an antipsychotic-like action through inhibition of PDE4 activity, indicating that it may be possible to treat psychiatric disorders by directly modulating PDE4-specific cAMP hydrolysis.
In animal models, treatment with PDE4 inhibitors has been found to enhance several models of learning and memory. This finding might indicate a use of PDE4 inhibitors as memory-enhancing agents and for decreasing the memory loss occurring in various types of neurodegenerative disease, in particular—but not limited to—Alzheimer's disease.
Safety-related issues are a significant challenge in the development of PDE4 inhibitors. Reported side effects or adverse events of PDE4 inhibitors include emesis, nausea, dyspepsia, diarrhea, abdominal pain, and headache. A potential side effect that has been observed in several animal models treated with PDE4 inhibitors, but has not been observed in humans yet, is arteritis. Arteritis is characterized by blood vessel inflammation, hemorrhage, and necrosis, and is considered irreversible in animals. The reported side effects of PDE4 inhibitors take place in the brain, gastrointestinal tract, arteries, or in the whole body due to systemic exposure to PDE4 inhibitors. In particular, one problem with PDE4 inhibitors is their tendency to promote emesis, which is mediated at least in part via actions in the central nervous system. In particular, PDE4 is present in parietal cells and emetic centers. A correlation has been found between the emetic effect of PDE4 inhibitors and the occupancy of the rolipram binding site of PDE4 by these inhibitors, indicating that such side effects are directly related to on-target activity. Hence, it will be appreciated that it is difficult to separate the effects on emesis from more desirable effects on the pathology to be treated.
Many development programs of PDE4 inhibitors have been discontinued due to side effects. The full potential of PDE4 inhibitors may not be realized as the administered dose is limited by side effects. Many dose-limiting side effects reflect an adverse interaction of the compound with PDE4 expressed in non-target tissues. In other words, adverse effects of PDE4 inhibitors may be directly related to on-target activity. Hence it will be appreciated that systemic exposure to PDE4 inhibitors should preferably be avoided if not required by the pathology to be treated.
Local application is a first possibility to reduce systemic exposure to a drug compound, by directly delivering the drug compound to the intended site of action and possibly reducing the quantity of drug compound that is required in order to observe a clinically significant effect. Despite the fact that direct local application is preferred in medical practice, there can still be concerns regarding drug levels reached into the systemic circulation. For example, the treatment of airway diseases by local delivery by for instance inhalation, poses the risk of systemic exposure due to large amounts entering the GI tract and/or systemic absorption through the lungs. For the treatment of eye diseases by local delivery, also significant amounts of compound can enter the GI tract and/or systemic circulation due to the low permeability of the cornea, low capacity for fluid, efficient drainage and presence of blood vessels in the eyes and eyelids. Also for dermal applications, local injections and implantable medical devices, there is a severe risk of leakage into the systemic circulation. Therefore, in addition to the physical local application, it is preferable that the compounds display additional chemical or biological properties that will minimize systemic exposure.
Soft drugs are biologically active compounds that are inactivated once they enter the systemic circulation. This inactivation can be achieved in the liver, but the preferred inactivation should occur in the blood. These compounds, once applied locally to the target tissue/organ exert their desired effect locally. When they leak out of this target tissue/organ into the systemic circulation, they are very rapidly inactivated. Thus, soft drugs of choice are sufficiently stable in the target tissue/organ to exert the desired biological effect, but are rapidly degraded in the blood to biologically inactive compounds. In addition, it is highly preferable that the soft drugs of choice have retention at their biological target. This property will limit the number of daily applications and is highly desired to reduce the total load of drug and metabolites and in addition will significantly increase the patient compliance.
In conclusion, there is a continuing need to design and develop soft PDE4 inhibitors for the treatment of a wide range of disease states. The compounds described herein and pharmaceutically acceptable compositions thereof are useful for treating or lessening the severity of a variety of disorders or conditions that can be treated via modulation of PDE4 activity.
More specifically, the compounds of the invention are preferably used in the prevention and/or treatment of at least one disease or disorder, in which the PDE4 family is involved, such as several inflammatory diseases or fibrotic diseases, which can be treated via local application of a drug compound. Such diseases include, but are not limited to idiopathic pulmonary fibrosis, inflammatory eye diseases such as dry eye disease or allergic eye disease; inflammatory airway diseases such as asthma or chronic obstructive pulmonary disease, skin diseases such as atopic dermatitis or psoriasis and intestinal diseases such as inflammatory bowel disease.